Neuronal loss, which has been considered to be directly linked to the major neurological manifestations of Alzheimer's disease (AD), is an important target for AD therapy although the pathological mechanism leading to neuronal loss still remains unknown. In vitro, a variety of AD-related insults, such as overexpression of FAD-related mutants and increased levels of toxic amyloid b peptides (Aβs) derived from Amyloid-β precursor protein (APP), induce neuronal cell death via multiple death pathways.
Familial AD (FAD) has been identified to be caused by missense mutations in three genes: APP, presenilin-1 (PS1), and presenilin-2 (PS2) [Shastry and Giblin, 1999]. Although it remains unknown how these mutant genes contribute to neuronal loss in FAD brain in vivo, multiple groups have provided evidence that expression of the FAD-associated mutant APP and PS genes causes neuronal cell death in cultured cells (Yamatsuji et al., 1996a, b; Wolozin et al., 1996; Zhao et al., 1997; Nishimura et al, 1998; Luo et al., 1999; Hashimoto et al., 2000) and primary cortical neurons (Niikura et al., 2004).
In addition, increased levels of toxic Aβs, which are considered to be closely linked to AD pathogenesis (Hardy and Selkoe, 2002), result in neuronal cell death in vitro although superphysiological concentrations of Aβ are required (Loo et al., 1994; Hashimoto et al., 2001; Hashimoto et al., 2004).
By performing a ‘death-trap’ screening, an unbiased functional screening of molecules that allows dying cells to survive, with a cDNA library constructed from an occipital lobe of the brain of an autopsy-diagnosed AD patient, we identified a cDNA encoding the 24 amino acids peptide MAPRGFSCLLLLTSEIDLPVKRRA (SEQ ID NO:1) named Humanin (HN) (WO01/021787) Polypeptide inhibiting neuronal cell death, Humanin), which suppresses neuronal cell death by all AD-related insults such as various FAD genes, anti-APP antibody, and neurotoxic Aβ peptides (Hashimoto et al., 2001a and b; Nishimoto et al., 2004). Ten mM of HN completely inhibits neurotoxicity induced by various AD-related insults. It has also been shown in follow-up studies that HN is also effective in inhibiting certain types of neuronal and non-neuronal cell death, such as serum-deprivation-induced cell death of PC12h cells (Kariya et al., 2002) and lymphocytes (Kariya et al., 2003), Aβ toxicity in human cerebrovascular smooth muscle cells (Jung et al, 2003), and prion-derived peptide-induced neurotoxicity (Sponne et al., 2004).
We have shown that HN is secreted from cells and inhibits neuronal cell death by AD-related insults from outside of cells via its putative receptor on the membrane (Hashimoto et al., 2001a; Nishimoto et al., 2004).
Most recently, Ying et al. (2004) have reported that HN inhibits Aβ (1-42)-induced neurotoxicity by binding to pertussis toxin (PTX)-sensitive G protein-coupled human formylpeptide receptor-like-1 (FPRL-1) as a HN receptor using PC12 neuroblastoma cells. They suggested that HN blocks Aβ-induced neurotoxicity by competing with Aβ for FPRL-1. However, after we have studied how FPRL-1 is involved in HN-mediated protection against AD-related neuronal insults, we found that FPRL-1 is not involved in HN-mediated neuroprotection in F11 neurohybrid cells or primary cortical neurons (Hashimoto et al., 2005), indicating that there are other receptors than FPRL-1, which may mediate HN-induced neuroprotection.
In addition, we revealed that STAT3 as well as a certain kind of tyrosine kinase are involved in HN-mediated neuroprotection (Hashimoto et al. 2005), suggesting that some cytokine receptor-like receptors are involved in their signaling pathway.
gp130 is a component of cytokine receptor common to interleukin-6 (IL-6) receptor family members. gp130-containing receptors are stimulated by several type I cytokines consisting of IL-6, IL-11, Leukemia-inhibitory factor (LIF), ciliary neurotrophic factor (CNTF), OncostatinM (OSM), and Cardiotropin-1. Binding of these cytokines to the above cognate receptors leads to homodimerization of gp130, or to heterodimerization between gp130 and a gp130-related receptor such as the LIF receptor, the OSM receptor or WSX-1 (IL-27 receptor), eventually transmitting cytokine signals to intracellular signal cascades mediated by both JAK/STAT and RAS/MAPK signaling pathways (Taga et al., 1997; Boulay et al., 2003; Boulanger et al., 2004). Most recently, it has been shown that IL-27 (IL-27p28/EBV-induced gene 3), which belongs to IL-6/IL-12 family cytokines, modifies Th-1 and Th-2 immunological response (Yoshida et al., 2004) by binding to WSX-1/gp130 (Plan et al, 2004). CNTF receptor alpha chain (CNTR-R) is a gp130-related receptor, which does not have an intracellular signaling domain. WO01/021787 and WO03/097687.